The long term goals of this project are to delineate the mechanisms of autoimmune disease. The murine experimental autoimmune thyroiditis model of an organ specific autoimmune disease will be used in this study. There are two specific aims in this proposal. 1. During organ specific autoimmune disease, self-tolerance is lost and autoantibodies to indiginous antigens arise through, as yet, an unknown mechanism. There are two possible pathways through which auto Ab may arise. In the germline immunoglobulin variable gene repertoire, there exist autoreactive V genes. Pathologic autoantibodies may be derived from these autoreactive, germline encoded V genes. Alternatively, autoreactive antibodies could arise from non-autoreactive germline V genes as a result of post- immunization somatic mutation. The first pathway could be explained totally by a loss of immunoregulation without the need of an antigen driven maturation process. The second pathway would necessitate an the presence of antigen to drive maturation, however, the process may be initiated by a unrelated antigen. We therefore, plan to clone, using recombinant DNA technology, the V genes of anti-mouse thyroglobulin (MTg) Ab developing during the disease process. The selection of V genes used will be compared to that of "natural" Ab that are autoreactive with MTg. The results from these experiments could point to one these pathways and give insight into the processes which induce autoimmune disease. 2. In the EAT model, disease can be easily induced by immunization with mouse thyroglobulin. The specific determinants involved in this induction process are not known. By chemical or enzymatic cleavage of MTg, we propose to define specific peptide sequences which are pathogenic or which can induce protective immunosuppression. Using the DNA probes derived from the amino acid sequence of these peptides, the location of these sites within the MTg molecule will be determined. Furthermore, we will attempt to distinguish determinants associated with development of disease from those which are involved in inducing autoantibody production alone. These finding would further our understanding of the nature of autoantigenic determinants.